ß-hydroxybutyrate administered at reperfusion reduces infarct size and preserves cardiac function by improving mitochondrial function through autophagy in male mice.

Ischemia/reperfusion (I/R) injury after revascularization contributes ∼50% of infarct size and causes heart failure, for which no established clinical treatment exists. ß-hydroxybutyrate (ß-OHB), which serves as both an energy source and a signaling molecule, has recently been reported to be cardioprotective when administered immediately before I/R and continuously after reperfusion. This study aims to determine whether administering ß-OHB at the time of reperfusion with a single dose can alleviate I/R injury and, if so, to define the mechanisms involved. We found plasma ß-OHB levels were elevated during ischemia in STEMI patients, albeit not to myocardial protection level, and decreased after revascularization. In mice, compared with normal saline, ß-OHB administrated at reperfusion reduced infarct size (by 50%) and preserved cardiac function, as well as activated autophagy and preserved mtDNA levels in the border zone. Our treatment with one dose ß-OHB reached a level achievable with fasting and strenuous physical activity. In neonatal rat ventricular myocytes (NRVMs) subjected to I/R, ß-OHB at physiologic level reduced cell death, increased autophagy, preserved mitochondrial mass, function, and membrane potential, in addition to attenuating reactive oxygen species (ROS) levels. ATG7 knockdown/knockout abolished the protective effects of ß-OHB observed both in vitro and in vivo. Mechanistically, ß-OHB's cardioprotective effects were associated with inhibition of mTOR signaling. In conclusion, ß-OHB, when administered at reperfusion, reduces infarct size and maintains mitochondrial homeostasis by increasing autophagic flux (potentially through mTOR inhibition). Since ß-OHB has been safely tested in heart failure patients, it may be a viable therapeutic to reduce infarct size in STEMI patients.

Role of the ST6GAL1 sialyltransferase in regulating ovarian cancer cell metabolism.

The ST6GAL1 sialyltransferase, which adds α2-6-linked sialic acids to N-glycosylated proteins, is upregulated in many malignancies including ovarian cancer. Through its activity in sialylating select surface receptors, ST6GAL1 modulates intracellular signaling to regulate tumor cell phenotype. ST6GAL1 has previously been shown to act as a survival factor that protects cancer cells from cytotoxic stressors such as hypoxia. In the present study, we investigated a role for ST6GAL1 in tumor cell metabolism. ST6GAL1 was overexpressed (OE) in OV4 ovarian cancer cells, which have low endogenous ST6GAL1, or knocked-down (KD) in ID8 ovarian cancer cells, which have high endogenous ST6GAL1. OV4 and ID8 cells with modulated ST6GAL1 expression were grown under normoxic or hypoxic conditions, and metabolism was assessed using Seahorse technology. Results showed that cells with high ST6GAL1 expression maintained a higher rate of oxidative metabolism than control cells following treatment with the hypoxia mimetic, desferrioxamine (DFO). This enrichment was not due to an increase in mitochondrial number. Glycolytic metabolism was also increased in OV4 and ID8 cells with high ST6GAL1 expression, and these cells displayed greater activity of the glycolytic enzymes, hexokinase and phosphofructokinase. Metabolism maps were generated from the combined Seahorse data, which suggested that ST6GAL1 functions to enhance the overall metabolism of tumor cells. Finally, we determined that OV4 and ID8 cells with high ST6GAL1 expression were more invasive under conditions of hypoxia. Collectively, these results highlight the importance of sialylation in regulating the metabolic phenotype of ovarian cancer cells.

Mitochondrial haplotype modulates genome expression and mitochondrial structure/function in cardiomyocytes following volume overload.

Mitochondrial DNA (mtDNA) haplotype regulates mitochondrial structure/function and reactive oxygen species in aortocaval fistula (ACF) in mice. Here, we unravel the mitochondrial haplotype effects on cardiomyocyte mitochondrial ultrastructure and transcriptome response to ACF in vivo. Phenotypic responses and quantitative transmission electron microscopy (TEM) and RNA sequence at 3 days were determined after sham surgery or ACF in vivo in cardiomyocytes from wild-type (WT) C57BL/6J (C57n:C57mt) and C3H/HeN (C3Hn:C3Hmt) and mitochondrial nuclear exchange mice (C57n:C3Hmt or C3Hn:C57mt). Quantitative TEM of cardiomyocyte mitochondria C3HWT hearts have more electron-dense compact mitochondrial cristae compared with C57WT. In response to ACF, mitochondrial area and cristae integrity are normal in C3HWT; however, there is mitochondrial swelling, cristae lysis, and disorganization in both C57WT and MNX hearts. Tissue analysis shows that C3HWT hearts have increased autophagy, antioxidant, and glucose fatty acid oxidation-related genes compared with C57WT. Comparative transcriptomic analysis of cardiomyocytes from ACF was dependent upon mtDNA haplotype. C57mtDNA haplotype was associated with increased inflammatory/protein synthesis pathways and downregulation of bioenergetic pathways, whereas C3HmtDNA showed upregulation of autophagy genes. In conclusion, ACF in vivo shows a protective response of C3Hmt haplotype that is in large part driven by mitochondrial nuclear genome interaction.NEW & NOTEWORTHY The results of this study support the effects of mtDNA haplotype on nuclear gene expression in cardiomyocytes. Currently, there is no acceptable therapy for volume overload due to mitral regurgitation. The findings of this study could suggest that mtDNA haplotype activates different pathways after ACF warrants further investigations on human population of heart disease from different ancestry backgrounds.

Scientific opportunities in resilience research for cardiovascular health and wellness. Report from a National Heart, Lung, and Blood Institute workshop.

Exposure of biological systems to acute or chronic insults triggers a host of molecular and physiological responses to either tolerate, adapt, or fully restore homeostasis; these responses constitute the hallmarks of resilience. Given the many facets, dimensions, and discipline-specific focus, gaining a shared understanding of "resilience" has been identified as a priority for supporting advances in cardiovascular health. This report is based on the working definition: "Resilience is the ability of living systems to successfully maintain or return to homeostasis in response to physical, molecular, individual, social, societal, or environmental stressors or challenges," developed after considering many factors contributing to cardiovascular resilience through deliberations of multidisciplinary experts convened by the National Heart, Lung, and Blood Institute during a workshop entitled: "Enhancing Resilience for Cardiovascular Health and Wellness." Some of the main emerging themes that support the possibility of enhancing resilience for cardiovascular health include optimal energy management and substrate diversity, a robust immune system that safeguards tissue homeostasis, and social and community support. The report also highlights existing research challenges, along with immediate and long-term opportunities for resilience research. Certain immediate opportunities identified are based on leveraging existing high-dimensional data from longitudinal clinical studies to identify vascular resilience measures, create a 'resilience index,' and adopt a life-course approach. Long-term opportunities include developing quantitative cell/organ/system/community models to identify resilience factors and mechanisms at these various levels, designing experimental and clinical interventions that specifically assess resilience, adopting global sharing of resilience-related data, and cross-domain training of next-generation researchers in this field.

Mito-Mendelian interactions alter in vivo glucose metabolism and insulin sensitivity in healthy mice.

The regulation of euglycemia is essential for human health with both chronic hypoglycemia and hyperglycemia having detrimental effects. It is well documented that the incidence of type 2 diabetes increases with age and exhibits racial disparity. Interestingly, mitochondrial DNA (mtDNA) damage also accumulates with age and its sequence varies with geographic maternal origins (maternal race). From these two observations, we hypothesized that mtDNA background may contribute to glucose metabolism and insulin sensitivity. Pronuclear transfer was used to generate mitochondrial-nuclear eXchange (MNX) mice to directly test this hypothesis, by assessing physiologic parameters of glucose metabolism in nuclear isogenic C57BL/6J mice harboring either a C57BL/6J (C57n:C57mt wild type-control) or C3H/HeN mtDNA (C57n:C3Hmt-MNX). All mice were fed normal chow diets. MNX mice were significantly leaner, had lower leptin levels, and were more insulin sensitive, with lower modified Homeostatic Model Assessment of Insulin Resistance (mHOMA-IR) values and enhanced insulin action when compared with their control counterparts. Further interrogation of muscle insulin signaling revealed higher phosphorylated Akt/total Akt ratios in MNX animals relative to control, consistent with greater insulin sensitivity. Overall, these results are consistent with the hypothesis that different mtDNA combinations on the same nuclear DNA (nDNA) background can significantly impact glucose metabolism and insulin sensitivity in healthy mice.NEW & NOTEWORTHY Different mitochondrial DNAs on the same nuclear genetic background can significantly impact body composition, glucose metabolism, and insulin sensitivity in healthy mice.

Unlocking the Secrets of Mitochondria in the Cardiovascular System: Path to a Cure in Heart Failure­A Report from the 2018 National Heart, Lung, and Blood Institute Workshop

Mitochondria have emerged as a central factor in the pathogenesis and progression of heart failure, and other cardiovascular diseases, as well, but no therapies are available to treat mitochondrial dysfunction. The National Heart, Lung, and Blood Institute convened a group of leading experts in heart failure, cardiovascular diseases, and mitochondria research in August 2018. These experts reviewed the current state of science and identified key gaps and opportunities in basic, translational, and clinical research focusing on the potential of mitochondria-based therapeutic strategies in heart failure. The workshop provided short- and long-term recommendations for moving the field toward clinical strategies for the prevention and treatment of heart failure and cardiovascular diseases by using mitochondria-based approaches.

Vascular Endothelial Mitochondrial Function Predicts Death or Pulmonary Outcomes in Preterm Infants.

RATIONALE: Vascular endothelial mitochondrial dysfunction contributes to the pathogenesis of several oxidant stress-associated disorders. Oxidant stress is a major contributor to the pathogenesis of bronchopulmonary dysplasia (BPD), a chronic lung disease of prematurity that often leads to sequelae in adult survivors. OBJECTIVES: This study was conducted to identify whether differences in mitochondrial bioenergetic function and oxidant generation in human umbilical vein endothelial cells (HUVECs) obtained from extremely preterm infants were associated with risk for BPD or death before 36 weeks postmenstrual age. METHODS: HUVEC oxygen consumption and superoxide and hydrogen peroxide generation were measured in 69 infants. MEASUREMENTS AND MAIN RESULTS: Compared with HUVECs from infants who survived without BPD, HUVECs obtained from infants who developed BPD or died had a lower maximal oxygen consumption rate (mean ± SEM, 107 ± 8 vs. 235 ± 22 pmol/min/30,000 cells; P < 0.001), produced more superoxide after exposure to hyperoxia (mean ± SEM, 89,807 ± 16,616 vs. 162,706 ± 25,321 MitoSOX Red fluorescence units; P < 0.05), and released more hydrogen peroxide into the supernatant after hyperoxia exposure (mean ± SEM, 1,879 ± 278 vs. 842 ± 119 resorufin arbitrary fluorescence units; P < 0.001). CONCLUSIONS: Our results indicating that endothelial cells of premature infants who later develop BPD or die have impaired mitochondrial bioenergetic capacity and produce more oxidants at birth suggest that the vascular endothelial mitochondrial dysfunction seen at birth in these infants persists through their postnatal life and contributes to adverse pulmonary outcomes and increased early mortality.

Mitochondrial DNA mutations and cardiovascular disease.

PURPOSE OF REVIEW: Cardiovascular disease (CVD) is responsible for more morbidity and mortality worldwide than any other ailment. Strategies for reducing CVD prevalence must involve identification of individuals at high risk for these diseases, and the prevention of its initial development. Such preventive efforts are currently limited by an incomplete understanding of the genetic determinants of CVD risk. In this review, evidence for the involvement of inherited mitochondrial mutations in development of CVD is examined. RECENT FINDINGS: Several forms of CVD have been documented in the presence of pathogenic mitochondrial DNA (mtDNA) mutations, both in isolation and as part of larger syndromes. Other 'natural' mtDNA polymorphisms not overtly tied to any pathology have also been associated with alterations in mitochondrial function and individual risk for CVD, but until very recently these studies have been merely correlative. Fortunately, novel animal models are now allowing investigators to define a causal relationship between inherited 'natural' mtDNA polymorphisms, and cardiovascular function and pathology. SUMMARY: Cardiovascular involvement is highly prevalent among patients with pathogenic mtDNA mutations. The relationship between CVD susceptibility and 'natural' mtDNA polymorphisms requires further investigation, but will be aided in the near future by several novel experimental models.

Mitochondrial DNA damage and vascular function in patients with diabetes mellitus and atherosclerotic cardiovascular disease.

OBJECTIVE: Prior studies demonstrate mitochondrial dysfunction with increased reactive oxygen species generation in peripheral blood mononuclear cells in diabetes mellitus. Oxidative stress-mediated damage to mitochondrial DNA promotes atherosclerosis in animal models. Thus, we evaluated the relation of mitochondrial DNA damage in peripheral blood mononuclear cells s with vascular function in patients with diabetes mellitus and with atherosclerotic cardiovascular disease. APPROACH AND RESULTS: We assessed non-invasive vascular function and mitochondrial DNA damage in 275 patients (age 57 ± 9 years, 60 % women) with atherosclerotic cardiovascular disease alone (N = 55), diabetes mellitus alone (N = 74), combined atherosclerotic cardiovascular disease and diabetes mellitus (N = 48), and controls age >45 without diabetes mellitus or atherosclerotic cardiovascular disease (N = 98). Mitochondrial DNA damage measured by quantitative PCR in peripheral blood mononuclear cells was higher with clinical atherosclerosis alone (0.55 ± 0.65), diabetes mellitus alone (0.65 ± 1.0), and combined clinical atherosclerosis and diabetes mellitus (0.89 ± 1.32) as compared to control subjects (0.23 ± 0.64, P < 0.0001). In multivariable models adjusting for age, sex, and relevant cardiovascular risk factors, clinical atherosclerosis and diabetes mellitus remained associated with higher mitochondrial DNA damage levels (ß = 0.14 ± 0.13, P = 0.04 and ß = 0.21 ± 0.13, P = 0.002, respectively). Higher mitochondrial DNA damage was associated with higher baseline pulse amplitude, a measure of arterial pulsatility, but not with flow-mediated dilation or hyperemic response, measures of vasodilator function. CONCLUSIONS: We found greater mitochondrial DNA damage in patients with diabetes mellitus and clinical atherosclerosis. The association of mitochondrial DNA damage and baseline pulse amplitude may suggest a link between mitochondrial dysfunction and excessive small artery pulsatility with potentially adverse microvascular impact.

Bioenergetic programming of macrophages by the apolipoprotein A-I mimetic peptide 4F.

The apoA-I (apolipoprotein A-I) mimetic peptide 4F favours the differentiation of human monocytes to an alternatively activated M2 phenotype. The goal of the present study was to test whether the 4F-mediated differentiation of MDMs (monocyte-derived macrophages) requires the induction of an oxidative metabolic programme. 4F treatment induced several genes in MDMs that play an important role in lipid metabolism, including PPARγ (peroxisome-proliferator-activated receptor γ) and CD36. Addition of 4F was associated with a significant increase in FA (fatty acid) uptake and oxidation compared with vehicle treatment. Mitochondrial respiration was assessed by measurement of the OCR (oxygen-consumption rate). 4F increased basal and ATP-linked OCR as well as maximal uncoupled mitochondrial respiration. These changes were associated with a significant increase in ΔΨm (mitochondrial membrane potential). The increase in metabolic activity in 4F-treated MDMs was attenuated by etomoxir, an inhibitor of mitochondrial FA uptake. Finally, addition of the PPARγ antagonist T0070907 to 4F-treated MDMs reduced the expression of CD163 and CD36, cell-surface markers for M2 macrophages, and reduced basal and ATP-linked OCR. These results support our hypothesis that the 4F-mediated differentiation of MDMs to an anti-inflammatory phenotype is due, in part, to an increase in FA uptake and mitochondrial oxidative metabolism.

Mitochondrial-nuclear genome interactions in non-alcoholic fatty liver disease in mice.

NAFLD (non-alcoholic fatty liver disease) involves significant changes in liver metabolism characterized by oxidative stress, lipid accumulation and fibrogenesis. Mitochondrial dysfunction and bioenergetic defects also contribute to NAFLD. In the present study, we examined whether differences in mtDNA influence NAFLD. To determine the role of mitochondrial and nuclear genomes in NAFLD, MNX (mitochondrial-nuclear exchange) mice were fed an atherogenic diet. MNX mice have mtDNA from C57BL/6J mice on a C3H/HeN nuclear background and vice versa. Results from MNX mice were compared with wild-type C57BL/6J and C3H/HeN mice fed a control or atherogenic diet. Mice with the C57BL/6J nuclear genome developed more macrosteatosis, inflammation and fibrosis compared with mice containing the C3H/HeN nuclear genome when fed the atherogenic diet. These changes were associated with parallel alterations in inflammation and fibrosis gene expression in wild-type mice, with intermediate responses in MNX mice. Mice with the C57BL/6J nuclear genome had increased State 4 respiration, whereas MNX mice had decreased State 3 respiration and RCR (respiratory control ratio) when fed the atherogenic diet. Complex IV activity and most mitochondrial biogenesis genes were increased in mice with the C57BL/6J nuclear or mitochondrial genome, or both fed the atherogenic diet. These results reveal new interactions between mitochondrial and nuclear genomes and support the concept that mtDNA influences mitochondrial function and metabolic pathways implicated in NAFLD.

The Bioenergetic Health Index: a new concept in mitochondrial translational research.

Bioenergetics has become central to our understanding of pathological mechanisms, the development of new therapeutic strategies and as a biomarker for disease progression in neurodegeneration, diabetes, cancer and cardiovascular disease. A key concept is that the mitochondrion can act as the 'canary in the coal mine' by serving as an early warning of bioenergetic crisis in patient populations. We propose that new clinical tests to monitor changes in bioenergetics in patient populations are needed to take advantage of the early and sensitive ability of bioenergetics to determine severity and progression in complex and multifactorial diseases. With the recent development of high-throughput assays to measure cellular energetic function in the small number of cells that can be isolated from human blood these clinical tests are now feasible. We have shown that the sequential addition of well-characterized inhibitors of oxidative phosphorylation allows a bioenergetic profile to be measured in cells isolated from normal or pathological samples. From these data we propose that a single value-the Bioenergetic Health Index (BHI)-can be calculated to represent the patient's composite mitochondrial profile for a selected cell type. In the present Hypothesis paper, we discuss how BHI could serve as a dynamic index of bioenergetic health and how it can be measured in platelets and leucocytes. We propose that, ultimately, BHI has the potential to be a new biomarker for assessing patient health with both prognostic and diagnostic value.

Mitochondrial genetic background modulates bioenergetics and susceptibility to acute cardiac volume overload.

Dysfunctional bioenergetics has emerged as a key feature in many chronic pathologies such as diabetes and cardiovascular disease. This has led to the mitochondrial paradigm in which it has been proposed that mtDNA sequence variation contributes to disease susceptibility. In the present study we show a novel animal model of mtDNA polymorphisms, the MNX (mitochondrial-nuclear exchange) mouse, in which the mtDNA from the C3H/HeN mouse has been inserted on to the C57/BL6 nuclear background and vice versa to test this concept. Our data show a major contribution of the C57/BL6 mtDNA to the susceptibility to the pathological stress of cardiac volume overload which is independent of the nuclear background. Mitochondria harbouring the C57/BL6J mtDNA generate more ROS (reactive oxygen species) and have a higher mitochondrial membrane potential relative to those with C3H/HeN mtDNA, independent of nuclear background. We propose this is the primary mechanism associated with increased bioenergetic dysfunction in response to volume overload. In summary, these studies support the 'mitochondrial paradigm' for the development of disease susceptibility, and show that the mtDNA modulates cellular bioenergetics, mitochondrial ROS generation and susceptibility to cardiac stress.

Methods for defining distinct bioenergetic profiles in platelets, lymphocytes, monocytes, and neutrophils, and the oxidative burst from human blood.

Peripheral blood mononuclear cells and platelets have long been recognized as having the potential to act as sensitive markers for mitochondrial dysfunction in a broad range of pathological conditions. However, the bioenergetic function of these cells has not been examined from the same donors, yet this is important for the selection of cell types for translational studies. Here, we demonstrate the measurement of cellular bioenergetics in isolated human monocytes, lymphocytes, and platelets, including the oxidative burst from neutrophils and monocytes from individual donors. With the exception of neutrophils, all cell types tested exhibited oxygen consumption that could be ascribed to oxidative phosphorylation with each having a distinct bioenergetic profile and distribution of respiratory chain proteins. In marked contrast, neutrophils were essentially unresponsive to mitochondrial respiratory inhibitors indicating that they have a minimal requirement for oxidative phosphorylation. In monocytes and neutrophils, we demonstrate the stimulation of the oxidative burst using phorbol 12-myristate 13-acetate and its validation in normal human subjects. Taken together, these data suggest that selection of cell type from blood cells is critical for assessing bioenergetic dysfunction and redox biology in translational research.

Beyond retrograde and anterograde signalling: mitochondrial-nuclear interactions as a means for evolutionary adaptation and contemporary disease susceptibility.

Although there is general agreement that most forms of common disease develop as a consequence of a combination of factors, including genetic, environmental and behavioural contributors, the actual mechanistic basis of how these factors initiate or promote diabetes, cancer, neurodegenerative and cardiovascular diseases in some individuals but not in others with seemingly identical risk factor profiles, is not clearly understood. In this respect, consideration of the potential role for mitochondrial genetics, damage and function in influencing common disease susceptibility seems merited, given that the prehistoric challenges were the original factors that moulded cellular function, and these were based upon the mitochondrial-nuclear relationships that were established during evolutionary history. These interactions were probably refined during prehistoric environmental selection events that, at present, are largely absent. Contemporary risk factors such as diet, sedentary lifestyle and increased longevity, which influence our susceptibility to a variety of chronic diseases were not part of the dynamics that defined the processes of mitochondrial-nuclear interaction, and thus cell function. Consequently, the prehistoric challenges that contributed to cell functionality and evolution should be considered when interpreting and designing experimental data and strategies. Although several molecular epidemiological studies have generally supported this notion, studies that probe beyond these associations are required. Such investigation will mark the initial steps for mechanistically addressing the provocative concept that contemporary human disease susceptibility is the result of prehistoric selection events for mitochondrial-nuclear function, which increased the probability for survival and reproductive success during evolution.

Convergent mechanisms for dysregulation of mitochondrial quality control in metabolic disease: implications for mitochondrial therapeutics.

Mitochondrial dysfunction is associated with a broad range of pathologies including diabetes, ethanol toxicity, metabolic syndrome and cardiac failure. It is now becoming clear that maintaining mitochondrial quality through a balance between biogenesis, reserve capacity and mitophagy is critical in determining the response to metabolic or xenobiotic stress. In diseases associated with metabolic stress, such as Type II diabetes and non-alcoholic and alcoholic steatosis, the mitochondria are subjected to multiple 'hits' such as hypoxia and oxidative and nitrative stress, which can overwhelm the mitochondrial quality control pathways. In addition, the underlying mitochondrial genetics that evolved to accommodate high-energy demand, low-calorie supply environments may now be maladapted to modern lifestyles (low-energy demand, high-calorie environments). The pro-oxidant and pro-inflammatory environment of a sedentary western lifestyle has been associated with modified redox cell signalling pathways such as steatosis, hypoxic signalling, inflammation and fibrosis. These data suggest that loss of mitochondrial quality control is intimately associated with the aberrant activation of redox cell signalling pathways under pathological conditions. In the present short review, we discuss evidence from alcoholic liver disease supporting this concept, the insights obtained from experimental models and the application of bioenergetic-based therapeutics in the context of maintaining mitochondrial quality.

Mitochondrial DNA Variations Modulate Alveolar Epithelial Mitochondrial Function and Oxidative Stress in Newborn Mice Exposed to Hyperoxia.

Oxidative stress is an important contributor to bronchopulmonary dysplasia (BPD), a form of chronic lung disease that is the most common morbidity in very preterm infants. Mitochondrial functional differences due to inherited and acquired mutations influence the pathogenesis of disorders in which oxidative stress plays a critical role. We previously showed using mitochondrial-nuclear exchange (MNX) mice that mitochondrial DNA (mtDNA) variations modulate hyperoxia-induced lung injury severity in a model of BPD. In this study, we studied the effects of mtDNA variations on mitochondrial function including mitophagy in alveolar epithelial cells (AT2) from MNX mice. We also investigated oxidant and inflammatory stress as well as transcriptomic profiles in lung tissue in mice and expression of proteins such as PINK1, Parkin and SIRT3 in infants with BPD. Our results indicate that AT2 from mice with C57 mtDNA had decreased mitochondrial bioenergetic function and inner membrane potential, increased mitochondrial membrane permeability and were exposed to higher levels of oxidant stress during hyperoxia compared to AT2 from mice with C3H mtDNA. Lungs from hyperoxia-exposed mice with C57 mtDNA also had higher levels of pro-inflammatory cytokines compared to lungs from mice with C3H mtDNA. We also noted changes in KEGG pathways related to inflammation, PPAR and glutamatergic signaling, and mitophagy in mice with certain mito-nuclear combinations but not others. Mitophagy was decreased by hyperoxia in all mice strains, but to a greater degree in AT2 and neonatal mice lung fibroblasts from hyperoxia-exposed mice with C57 mtDNA compared to C3H mtDNA. Finally, mtDNA haplogroups vary with ethnicity, and Black infants with BPD had lower levels of PINK1, Parkin and SIRT3 expression in HUVEC at birth and tracheal aspirates at 28 days of life when compared to White infants with BPD. These results indicate that predisposition to neonatal lung injury may be modulated by variations in mtDNA and mito-nuclear interactions need to be investigated to discover novel pathogenic mechanisms for BPD.

Mitochondrial Transfer to Host Cells from Ex Vivo Expanded Donor Hematopoietic Stem Cells.

Mitochondrial dysfunction is observed in various conditions, from metabolic syndromes to mitochondrial diseases. Moreover, mitochondrial DNA (mtDNA) transfer is an emerging mechanism that enables the restoration of mitochondrial function in damaged cells. Hence, developing a technology that facilitates the transfer of mtDNA can be a promising strategy for the treatment of these conditions. Here, we utilized an ex vivo culture of mouse hematopoietic stem cells (HSCs) and succeeded in expanding the HSCs efficiently. Upon transplantation, sufficient donor HSC engraftment was attained in-host. To assess the mitochondrial transfer via donor HSCs, we used mitochondrial-nuclear exchange (MNX) mice with nuclei from C57BL/6J and mitochondria from the C3H/HeN strain. Cells from MNX mice have C57BL/6J immunophenotype and C3H/HeN mtDNA, which is known to confer a higher stress resistance to mitochondria. Ex vivo expanded MNX HSCs were transplanted into irradiated C57BL/6J mice and the analyses were performed at six weeks post transplantation. We observed high engraftment of the donor cells in the bone marrow. We also found that HSCs from the MNX mice could transfer mtDNA to the host cells. This work highlights the utility of ex vivo expanded HSC to achieve the mitochondrial transfer from donor to host in the transplant setting.